Methylotrophic yeasts are those yeasts that are able to utilize methanol as a sole source of carbon and energy. Species of yeasts that have the biochemical pathways necessary for methanol utilization are classified in four genera, Hansenula, Pichia, Candida, and Torulopsis. These genera are somewhat artificial, having been based on cell morphology and growth characteristics, and do not reflect close genetic relationships (Billon-Grand, Mycotaxon 35:201-204, 1989; Kurtzman, Mycologia 84:72-76, 1992). Furthermore, not all species within these genera are capable of utilizing methanol as a source of carbon and energy. As a consequence of this classification, there are great differences in physiology and metabolism between individual species of a genus.
Methylotrophic yeasts are attractive candidates for use in recombinant protein production systems for several reasons. First, some methylotrophic yeasts have been shown to grow rapidly to high biomass on minimal defined media. Second, recombinant expression cassettes are genomically integrated and therefore mitotically stable. Third, these yeasts are capable of secreting large amounts of recombinant proteins. See, for example, Faber et al., Yeast 11:1331, 1995; Romanos et al., Yeast 8:423, 1992; Cregg et al., Bio/Technology 11:905, 1993; U.S. Pat. No. 4,855,242; U.S. Pat. No. 4,857,467; U.S. Pat. No. 4,879,231; and U.S. Pat. No. 4,929,555; and Raymond, U.S. Pat. Nos. 5,716,808, 5,736,383, 5,854,039, and 5,888,768.
In the commercial production of proteins via recombinant DNA technologies, it is often advantageous for the desired protein of interest to be secreted into the growth medium. Secretion of proteins from cells is generally accomplished by the presence of a short stretch of hydrophobic amino acids constituting the amino-terminal end of the translational product. This hydrophobic stretch is call the “secretory signal sequence,” and it is possible to use signal sequences to effect the secretion of heterologous proteins. This is generally accomplished by the construction of an DNA construct comprising a DNA sequence encoding a secretory signal sequence, into which a gene encoding the desired heterologous protein is inserted. When such a plasmid is transformed into a host cell, the host cell will express and secrete the desired protein into the growth medium.
At present, the only mode of achieving secretion of a heterologous protein product in Pichia methanolica is by way of a foreign secretory signal peptide. Because foreign gene's are not native to Pichia methanolica, the levels of heterologous protein expression are likely suboptimal as compared to a DNA construct incorporating a secretory signal sequence native to Pichia methanolica. 
Thus, there remains a need in the art to identify a secretory signal sequence native to Pichia methanolica to enable the use of methylotrophic yeasts for production of polypeptides of economic importance, including industrial enzymes and pharmaceutical proteins. The present invention provides such materials and methods as well as other, related advantages.